U.S. patent application number 10/400604 was filed with the patent office on 2003-10-02 for microscope having apparatus for determining the light power level of an illuminating light beam.
This patent application is currently assigned to Leica Microsystems Heidelberg GmbH. Invention is credited to Hay, William C..
Application Number | 20030184857 10/400604 |
Document ID | / |
Family ID | 7969570 |
Filed Date | 2003-10-02 |
United States Patent
Application |
20030184857 |
Kind Code |
A1 |
Hay, William C. |
October 2, 2003 |
Microscope having apparatus for determining the light power level
of an illuminating light beam
Abstract
A microscope has a light source that emits an illuminating light
beam for illumination of a specimen, an apparatus for determining
the light power level of the illuminating light beam and a beam
splitter separating measuring light out of the illuminating light
beam. The microscope permits determination of the light power level
of the illuminating light beam with an apparatus for simultaneous
color-selective detection of the measuring light.
Inventors: |
Hay, William C.;
(Heppenheim, DE) |
Correspondence
Address: |
DAVIDSON, DAVIDSON & KAPPEL, LLC
485 SEVENTH AVENUE, 14TH FLOOR
NEW YORK
NY
10018
US
|
Assignee: |
Leica Microsystems Heidelberg
GmbH
Mannheim
DE
|
Family ID: |
7969570 |
Appl. No.: |
10/400604 |
Filed: |
March 27, 2003 |
Current U.S.
Class: |
359/385 ;
359/368; 359/380; 359/431; 359/489.09; 359/489.19; 359/589;
359/634 |
Current CPC
Class: |
G02B 21/06 20130101;
G02B 21/18 20130101 |
Class at
Publication: |
359/385 ;
359/368; 359/380; 359/431; 359/498; 359/589; 359/634 |
International
Class: |
G02B 021/00; G02B
021/06; G02B 023/00; G02B 005/30; G02B 027/28; G02B 005/28; G02B
027/14 |
Foreign Application Data
Date |
Code |
Application Number |
Mar 30, 2002 |
DE |
DE 202 05 081.5 |
Claims
What is claimed is:
1. A microscope comprising: a light source that emits an
illuminating light beam for illumination of a specimen, a beam
splitter separating measuring light out of the illuminating light
beam, and an apparatus for determining the light power level of the
illuminating light beam, which receives the measuring light and
which comprises an apparatus for simultaneous color-selective
detection of the measuring light.
2. The microscope as defined in claim 1, wherein the apparatus for
simultaneous color-selective detection comprises a spatially
spectrally dividing element.
3. The microscope as defined in claim 2, wherein the spatially
spectrally dividing element is a prism.
4. The microscope as defined in claim 3, wherein one surface of the
prism constitutes the beam splitter.
5. The microscope as defined in claim 4, wherein the surface of the
prism is coated in partially reflective fashion.
6. The microscope as defined in claim 4, wherein at least one
surface of the prism is roughened.
7. The microscope as defined in claim 1, wherein the apparatus for
simultaneous color-selective detection comprises at least one
detector that receives the measuring light.
8. The microscope as defined in claim 7, wherein the detector
consists of several individual detectors that each receive
spectrally different components of the measuring light.
9. The microscope as defined in claim 7, wherein the detector
contains a photodiode or a photomultiplier or a photodiode row or a
photodiode array or a CCD element or a photomultiplier array or a
photomultiplier row.
10. The microscope as defined in claim 7, wherein the detector is
arranged directly on the apparatus for simultaneous color-selective
detection.
11. An apparatus for determining the light power level of an
illuminating light beam comprising: a beam splitter that separates
measuring light out of the illuminating light beam, and an
apparatus for simultaneous color-selective detection of the
measuring light.
12. The apparatus as defined in claim 11, wherein the apparatus for
simultaneous color-selective detection comprises a spatially
spectrally dividing element.
13. The apparatus as defined in claim 12, wherein the spatially
spectrally dividing element is a prism.
14. The apparatus as defined in claim 13, wherein one surface of
the prism constitutes the beam splitter.
15. The apparatus as defined in claim 14, wherein the surface of
the prism is coated in partially reflective fashion.
16. The apparatus as defined in claim 14, wherein at least one
surface of the prism is roughened.
17. The apparatus as defined in claim 11, wherein the apparatus for
simultaneous color-selective detection comprises at least one
detector that receives the measuring light.
18. The apparatus as defined in claim 17, wherein the detector
comprises several individual detectors that each receive spectrally
different components of the measuring light.
19. The apparatus as defined in claim 17, wherein the detector
contains a photodiode or a photomultiplier or a photodiode row or a
photodiode array or a CCD element or a photomultiplier array or a
photomultiplier row.
20. The apparatus as defined in claim 17, wherein the detector is
arranged directly on the apparatus for simultaneous color-selective
detection.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority to German utility model
application 202 05 081.5, which is hereby incorporated by reference
herein.
BACKGROUND
[0002] The invention concerns a microscope.
[0003] The invention further concerns an apparatus for determining
the light power level of an illuminating light beam.
[0004] It is general practice, in order to measure the power level
of a light beam, to divide a measuring beam out of the light beam
using a beam splitter and, by means of a detector that generates an
electrical signal proportional to the power level of the measuring
beam, firstly to determine the power level of the measuring beam
and then, with a knowledge of the splitting ratio of the beam
splitter, to draw conclusions as to the power level of the light
beam.
[0005] In scanning microscopy, a specimen is illuminated with a
light beam in order to observe the detected light emitted, as
reflected or fluorescent light, from the specimen. The illuminating
light beam can contain light of several wavelengths, for example in
order to excite several dyes simultaneously. The focus of an
illuminating light beam is moved in a specimen plane by means of a
controllable beam deflection device, generally by tilting two
mirrors; the deflection axes are usually perpendicular to one
another, so that one mirror deflects in the X direction and the
other in the Y direction. Tilting of the mirrors is brought about,
for example, by means of galvanometer positioning elements. The
power level of the detected light coming from the specimen is
measured as a function of the position of the scanning beam. The
positioning elements are usually equipped with sensors to ascertain
the present mirror position.
[0006] In confocal scanning microscopy specifically, a specimen is
scanned in three dimensions with the focus of a light beam. A
confocal scanning microscope generally comprises a light source, a
focusing optical system with which the light of the source is
focused onto a diaphragm (called the "excitation pinhole"), a beam
splitter, a beam deflection device for beam control, a microscope
optical system, a detection pinhole, and the detectors for
detecting the detected or fluorescent light. The illuminating light
is coupled in via a beam splitter. The fluorescent or reflected
light coming from the specimen travels back through the beam
deflection device to the beam splitter, passes through it, and is
then focused onto the detection pinhole behind which the detectors
are located. This detection arrangement is called a "descan"
arrangement. Detected light that does not derive directly from the
focus region takes a different light path and does not pass through
the detection pinhole, so that a point datum is obtained which
results, by sequential scanning of the specimen, in a
three-dimensional image. A three-dimensional image is usually
achieved by acquiring image data in layers. Commercial scanning
microscopes usually comprise a scanning module that is
flange-mounted onto the stand of a conventional light microscope,
the scanning module containing all the aforesaid elements
additionally necessary for scanning a specimen.
[0007] One known method of compensating for or correcting
fluctuations in the illuminating light power level is based on
dividing a measuring beam out of the illuminating light beam with a
beam splitter, and using the ratio of the measured power levels of
the measuring beam and detected light beam for image generation or
image calculation. This procedure is disclosed, for example, in the
publication by G. J. Brakenhoff, Journal of Microscopy, Vol. 117,
pt. Nov. 2, 1979, pp. 233-242.
[0008] German Unexamined Application DE 197 02 753 A1 discloses an
arrangement for monitoring the laser radiation coupled into a
scanning head, by means of a detection element onto which a portion
of the incoupled radiation is directed via a beam splitter. This
arrangement can be equipped with exchangeable filters in order to
make possible wavelength-dependent monitoring of the laser
radiation. The arrangement has the disadvantage that without a
filter wheel, only the overall power level of the illuminating
light beam, which in many applications is made up of light of
several wavelengths, can be monitored. For example, if the light of
two different lasers is combined into one illuminating light beam,
any fluctuation that occurs in the power level of a laser can be
recorded, but it cannot be corrected or compensated for because
allocation is not possible. The use of the filters solves this
problem, but with a great deal of outlay, especially in terms of
time, on the part of the microscope user. In addition, exchanging
of the filters by the user entails the risk of unintentional
misalignments and other external disruptions, in particular
vibrations and thermal influences, which limit the reproducibility
and accuracy of the measurement.
SUMMARY OF THE INVENTION
[0009] It is therefore an object of the invention to provide a
microscope that makes possible efficient, reliable, and accurate
determination and monitoring of the light power level of the
illuminating light beam even in multi-color applications.
[0010] The invention provides a microscope comprising:
[0011] a light source that emits an illuminating light beam for
illumination of a specimen,
[0012] a beam splitter separating measuring light out of the
illuminating light beam, and
[0013] an apparatus for determining the light power level of the
illuminating light beam,
[0014] which receives the measuring light and which comprises an
apparatus for simultaneous color-selective detection of the
measuring light.
[0015] A further object of the invention is to provide an apparatus
that allows efficiently and reliably to determine the light power
level of a multi-color illuminating light beam.
[0016] The present invention also provides an apparatus for
determining the light power level of an illuminating light beam
comprising: a beam splitter that separates measuring light out of
the illuminating light beam, and an apparatus for simultaneous
color-selective detection of the measuring light.
[0017] The invention has the advantage it makes possible reliable
and rapid (online) measurement and monitoring of the power level of
all spectral components of the illuminating light beam, and thus at
the same time creates the prerequisites for compensating, even in
the context of multi-color applications, for fluctuations in the
illuminating light power level of the illuminating light beam or
fluctuations in components of the illuminating light beam, for
example in a closed-loop control system.
[0018] In a preferred embodiment, the apparatus for simultaneous
color-selective detection comprises a spatially spectrally dividing
element that preferably is embodied as a prism. It can also be
embodied, for example, as a grating, in particular as a
transmission grating. In a very particularly preferred embodiment,
one surface (preferably coated) of the prism constitutes the beam
splitter. The coating can be, for example, a partial mirror
coating.
[0019] A further disadvantage exhibited by the arrangement known
from the existing art, namely that interference effects brought
about by multiple reflections occur at a plane-parallel beam
splitter substrate because of its thinness, and result in severe
intensity fluctuations at the detector, is also eliminated in the
apparatus according to the present invention. Since there are no
parallel surfaces in a prism, multiple reflections cannot in this
case result in interference phenomena at the detector.
[0020] In order to avoid reflections that, after multiple
deflection within the prism, ultimately strike the detector and
distort the signal there, individual surfaces of the prism can be
roughened.
[0021] In a preferred embodiment, the apparatus for simultaneous
color-selective detection comprises at least one detector that
receives the measured light. In a very particularly preferred
embodiment, the detector is made up of several individual detectors
that each receive spectrally different components of the measured
light. The detector can contain, for example, a photodiode or a
photomultiplier or a photodiode row or a photodiode array or a CCD
element or a photomultiplier array or a photomultiplier row. The
individual detectors are preferably each individually calibrated
for the wavelength that they receive.
[0022] In another preferred embodiment, the detector is arranged
directly on the apparatus for simultaneous color-selective
detection. Preferably the entry window of the detector is cemented
directly onto the exit surface of the prism. This prevents
disruptive interference effects due to multiple reflections between
the prism and detector.
BRIEF DESCRIPTION OF THE DRAWINGS
[0023] The subject matter of the invention is depicted in the
drawings and will be described below with reference to the Figures,
identically functioning elements being labeled with the same
reference characters. In the drawings:
[0024] FIG. 1 shows an apparatus according to the present invention
for determining the light output of an illuminating light beam;
[0025] FIG. 2 shows a further apparatus according to the present
invention for determining the light output of an illuminating light
beam;
[0026] FIG. 3 shows a further apparatus according to the present
invention for determining the light output of an illuminating light
beam;
[0027] FIG. 4 shows a further apparatus according to the present
invention for determining the light output of an illuminating light
beam; and
[0028] FIG. 5 shows a microscope according to the present
invention.
DETAILED DESCRIPTION
[0029] FIG. 1 shows an apparatus according to the present invention
for determining the light power level of an illuminating light
beam. The apparatus comprises a spatially spectrally dividing
element 1 that is embodied as prism 3. A first surface of the prism
is provided with a partially reflective coating 5, and constitutes
a beam splitter 7. Illuminating light beam 9 strikes beam splitter
7. At coating 5, 5% of the illuminating light beam is divided out
as measured light 11, which is spatially spectrally spread out by
prism 3 and leaves prism 3 through an exit surface 13. The measured
light then strikes a detector 15 that is embodied as photodiode row
17. In the individual detectors of the photodiode row, electrical
signals proportional in current intensity to the light power level
of the respective spectral component are generated.
[0030] FIG. 2 shows a further apparatus according to the present
invention for determining the light power level of an illuminating
light beam. In this embodiment the detector is cemented directly
onto the exit surface of the prism.
[0031] FIG. 3 shows a further apparatus according to the present
invention for determining the light power level of an illuminating
light beam. In this embodiment, a third surface 19 of prism 3 is
roughened in order to suppress unwanted reflections.
[0032] FIG. 4 shows a further apparatus according to the present
invention for determining the power level of an illuminating light
beam, which corresponds largely to the embodiment shown in FIG. 2.
In this embodiment, a third surface 19 of prism 3, and exit surface
13, are roughened in order to suppress undesired reflections.
[0033] FIG. 5 schematically shows a microscope 33 according to the
present invention, which is embodied as a confocal scanning
microscope. Light beam 37 coming from an illumination system 35 is
transported via a glass fiber 39 and, after being coupled out of
glass fiber 39 by means of optical system 41, strikes an apparatus
43 for determining the power level of illuminating light beam 37,
which corresponds largely to the apparatus shown in FIG. 1 having a
prism 3 and a photodiode row 17. Detector 15 generates electrical
signals proportional to the power level of the respective spectral
components of measured light 11, which are forwarded via conductor
45 to processing unit 47. By way of a beam splitter 49,
illuminating light beam 37 arrives at gimbal-mounted scanning
mirror 51 that guides the beam, through scanning optical system 53,
tube optical system 55, and objective 57, over or through specimen
59. In the case of non-transparent specimens 59, illuminating light
beam 37 is guided over the specimen surface. With biological
specimens 59 (preparations) or transparent specimens, illuminating
light beam 37 can also be guided through specimen 59. This means
that different focal planes of the specimen are successively
scanned by illuminating light beam 37. Subsequent assembly then
yields a three-dimensional image of the specimen. Detected light 61
proceeding from specimen 59 travels through objective 57, tube
optical system 55, and scanning optical system 53, and via scanning
mirror 51 to beam splitter 49, passes through the latter and
strikes a detector apparatus 63, which is embodied as a multi-band
detector. In detector apparatus 63, which is embodied as a
multi-band detector, electrical detected signals proportional to
the power level of the detected light are generated in spectrally
selective fashion and are forwarded via conductor 65 to processing
unit 47. In processing unit 47, the incoming analog signals are
first digitized and then digitally correlated with one another, and
corrected detected light power levels are determined. These are
forwarded to a PC 67. The corrected detected light power levels are
allocated, on the basis of a position signal of the gimbal-mounted
mirror, to the position of the associated grid point, and the data
of all the grid points are assembled into an image of specimen 69
that is displayed on a display 71. Illumination pinhole 73 and
detection pinhole 75 that are usually provided in a confocal
scanning microscope are schematically drawn in for the sake of
completeness. Omitted in the interest of better clarity, however,
are certain optical elements for guiding and shaping the light
beams. These are sufficiently familiar to a person skilled in this
art.
[0034] The invention has been described with reference to a
particular exemplary embodiment. It is self-evident, however, that
changes and modifications can be made without thereby leaving the
range of protection of the claims below.
* * * * *